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In general, females present with stronger immune responses than males, but scarce data are available on sex-specific differences in immunometabolism. In this study, we characterized porcine peripheral blood mononuclear cell (PBMC) and granulocyte energy metabolism using a Bayesian 13 C-metabolic flux analysis, which allowed precise determination of the glycolytic, pentose phosphate pathway (PPP), and tricarboxylic acid cycle (TCA) fluxes, together with an assessment of the superoxide anion radical (O 2 •- ) production and mitochondrial O 2 consumption. A principal component analysis allowed for identifying the cell type-specific patterns of metabolic plasticity. PBMCs displayed higher TCA cycle activity, especially glutamine-derived aspartate biosynthesis, which was directly related to mitochondrial respiratory activity and inversely related to O 2 •- production. In contrast, the granulocytes mainly utilized glucose via glycolysis, which was coupled to oxidative PPP utilization and O 2 •- production rates. The granulocytes of the males had higher oxidative PPP fluxes compared to the females, while the PBMCs of the females displayed higher non-oxidative PPP fluxes compared to the males associated with the T helper cell (CD3 + CD4 + ) subpopulation of PBMCs. The observed sex-specific differences were not directly attributable to sex steroid plasma levels, but we detected an inverse correlation between testosterone and aldosterone plasma levels and showed that aldosterone levels were related with non-oxidative PPP fluxes of both cell types.

Introduction: Supplementation with increased inspired oxygen fractions has been suggested to alleviate the harmful effects of tissue hypoxia during hemorrhagic shock (HS) and traumatic brain injury. However, the utility of therapeutic hyperoxia in critical care is disputed to this day as controversial evidence is available regarding its efficacy. Furthermore, in contrast to its hypoxic counterpart, the effect of hyperoxia on the metabolism of circulating immune cells remains ambiguous. Both stimulating and detrimental effects are possible; the former by providing necessary oxygen supply, the latter by generation of excessive amounts of reactive oxygen species (ROS). To uncover the potential impact of increased oxygen fractions on circulating immune cells during intensive care, we have performed a 13 C-metabolic flux analysis (MFA) on PBMCs and granulocytes isolated from two long-term, resuscitated models of combined acute subdural hematoma (ASDH) and HS in pigs with and without cardiovascular comorbidity., Methods: Swine underwent resuscitation after 2 h of ASDH and HS up to a maximum of 48 h after HS. Animals received normoxemia (P a O 2 = 80 - 120 mmHg) or targeted hyperoxemia (P a O 2 = 200 - 250 mmHg for 24 h after treatment initiation, thereafter P a O 2 as in the control group). Blood was drawn at time points T1 = after instrumentation, T2 = 24 h post ASDH and HS, and T3 = 48 h post ASDH and HS. PBMCs and granulocytes were isolated from whole blood to perform electron spin resonance spectroscopy, high resolution respirometry and 13 C-MFA. For the latter, we utilized a parallel tracer approach with 1,2- 13 C 2 glucose, U- 13 C glucose, and U- 13 C glutamine, which covered essential pathways of glucose and glutamine metabolism and supplied redundant data for robust Bayesian estimation. Gas chromatography-mass spectrometry further provided multiple fragments of metabolites which yielded additional labeling information. We obtained precise estimations of the fluxes, their joint credibility intervals, and their relations, and characterized common metabolic patterns with principal component analysis (PCA)., Results: 13 C-MFA indicated a hyperoxia-mediated reduction in tricarboxylic acid (TCA) cycle activity in circulating granulocytes which encompassed fluxes of glutamine uptake, TCA cycle, and oxaloacetate/aspartate supply for biosynthetic processes. We further detected elevated superoxide levels in the swine strain characterized by a hypercholesterolemic phenotype. PCA revealed cell type-specific behavioral patterns of metabolic adaptation in response to ASDH and HS that acted irrespective of swine strains or treatment group., Conclusion: In a model of resuscitated porcine ASDH and HS, we saw that ventilation with increased inspiratory O 2 concentrations (P a O 2 = 200 - 250 mmHg for 24 h after treatment initiation) did not impact mitochondrial respiration of PBMCs or granulocytes. However, Bayesian 13 C-MFA results indicated a reduction in TCA cycle activity in granulocytes compared to cells exposed to normoxemia in the same time period. This change in metabolism did not seem to affect granulocytes' ability to perform phagocytosis or produce superoxide radicals., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Wolfschmitt, Vogt, Hogg, Wachter, Stadler, Kapapa, Datzmann, Messerer, Hoffmann, Gröger, Münz, Mathieu, Mayer, Merz, Asfar, Calzia, Radermacher and Zink.)

The pentose phosphate pathway (PPP) plays a key role in the cellular regulation of immune function; however, little is known about the interplay of metabolic adjustments in granulocytes, especially regarding the non-oxidative PPP. For the determination of metabolic mechanisms within glucose metabolism, we propose a novel set of measures for 13 C-metabolic flux analysis based on ex vivo parallel tracer experiments ([1,2- 13 C]glucose, [U- 13 C]glucose, [4,5,6- 13 C]glucose) and gas chromatography-mass spectrometry labeling measurements of intracellular metabolites, such as sugar phosphates and their fragments. A detailed constraint analysis showed that the permission range for net and irreversible fluxes was limited to a three-dimensional space. The overall workflow, including its Bayesian flux estimation, resulted in precise flux distributions and pairwise confidence intervals, some of which could be represented as a line due to the strength of their correlation. The principal component analysis that was enabled by these behaviors comprised three components that explained 99.6% of the data variance. It showed that phagocytic stimulation reversed the direction of non-oxidative PPP net fluxes from ribose-5-phosphate biosynthesis toward glycolytic pathways. This process was closely associated with the up-regulation of the oxidative PPP to promote the oxidative burst.

Introduction: Sodium thiosulfate (Na 2 S 2 O 3 ), an H 2 S releasing agent, was shown to be organ-protective in experimental hemorrhage. Systemic inflammation activates immune cells, which in turn show cell type-specific metabolic plasticity with modifications of mitochondrial respiratory activity. Since H 2 S can dose-dependently stimulate or inhibit mitochondrial respiration, we investigated the effect of Na 2 S 2 O 3 on immune cell metabolism in a blinded, randomized, controlled, long-term, porcine model of hemorrhage and resuscitation. For this purpose, we developed a Bayesian sampling-based model for 13 C isotope metabolic flux analysis (MFA) utilizing 1,2- 13 C 2 -labeled glucose, 13 C 6 -labeled glucose, and 13 C 5 -labeled glutamine tracers., Methods: After 3 h of hemorrhage, anesthetized and surgically instrumented swine underwent resuscitation up to a maximum of 68 h. At 2 h of shock, animals randomly received vehicle or Na 2 S 2 O 3 (25 mg/kg/h for 2 h, thereafter 100 mg/kg/h until 24 h after shock). At three time points (prior to shock, 24 h post shock and 64 h post shock) peripheral blood mononuclear cells (PBMCs) and granulocytes were isolated from whole blood, and cells were investigated regarding mitochondrial oxygen consumption (high resolution respirometry), reactive oxygen species production (electron spin resonance) and fluxes within the metabolic network (stable isotope-based MFA)., Results: PBMCs showed significantly higher mitochondrial O 2 uptake and lower O 2 • - production in comparison to granulocytes. We found that in response to Na 2 S 2 O 3 administration, PBMCs but not granulocytes had an increased mitochondrial oxygen consumption combined with a transient reduction of the citrate synthase flux and an increase of acetyl-CoA channeled into other compartments, e.g., for lipid biogenesis., Conclusion: In a porcine model of hemorrhage and resuscitation, Na 2 S 2 O 3 administration led to increased mitochondrial oxygen consumption combined with stimulation of lipid biogenesis in PBMCs. In contrast, granulocytes remained unaffected. Granulocytes, on the other hand, remained unaffected. O 2 • - concentration in whole blood remained constant during shock and resuscitation, indicating a sufficient anti-oxidative capacity. Overall, our MFA model seems to be is a promising approach for investigating immunometabolism; especially when combined with complementary methods., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. The reviewer JB declared a past collaboration with the authors E-MW, MG and MH to the handling editor., (Copyright © 2023 Wolfschmitt, Hogg, Vogt, Zink, Wachter, Hezel, Zhang, Hoffmann, Gröger, Hartmann, Gässler, Datzmann, Merz, Hellmann, Kranz, Calzia, Radermacher and Messerer.)

Introduction: We previously showed that attenuated glucocorticoid receptor (GR) function in mice (GR dim/dim ) aggravates systemic hypotension and impairs organ function during endotoxic shock. Hemorrhagic shock (HS) causes impaired organ perfusion, which leads to tissue hypoxia and inflammation with risk of organ failure. Lung co-morbidities like chronic obstructive pulmonary disease (COPD) can aggravate tissue hypoxia via alveolar hypoxia. The most common cause for COPD is cigarette smoke (CS) exposure. Therefore, we hypothesized that affecting GR function in mice (GR dim/dim ) and pre-traumatic CS exposure would further impair hemodynamic stability and organ function after HS., Methods: After 3 weeks of CS exposure, anesthetized and mechanically ventilated GR dim/dim and GR +/+ mice underwent pressure-controlled HS for 1h via blood withdrawal (mean arterial pressure (MAP) 35mmHg), followed by 4h of resuscitation with re-transfusion of shed blood, colloid fluid infusion and, if necessary, continuous intravenous norepinephrine. Acid-base status and organ function were assessed together with metabolic pathways. Blood and organs were collected at the end of the experiment for analysis of cytokines, corticosterone level, and mitochondrial respiratory capacity. Data is presented as median and interquartile range., Results: Nor CS exposure neither attenuated GR function affected survival. Non-CS GR dim/dim mice had a higher need of norepinephrine to keep target hemodynamics compared to GR +/+ mice. In contrast, after CS exposure norepinephrine need did not differ significantly between GR dim/dim and GR +/+ mice. Non-CS GR dim/dim mice presented with a lower pH and increased blood lactate levels compared to GR +/+ mice, but not CS exposed mice. Also, higher plasma concentrations of some pro-inflammatory cytokines were observed in non-CS GR dim/dim compared to GR +/+ mice, but not in the CS group. With regards to metabolic measurements, CS exposure led to an increased lipolysis in GR dim/dim compared to GR +/+ mice, but not in non-CS exposed animals., Conclusion: Whether less metabolic acidosis or increased lipolysis is the reason or the consequence for the trend towards lower catecholamine need in CS exposed GR dim/dim mice warrants further investigation., Competing Interests: Authors CT, MK, and JB are employed by Boehringer Ingelheim Pharma GmbH & Co KG. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Wepler, Preuss, Tilp, Keck, Blender, Wachter, Merz, Vogt, Kress, Gröger, Hoffmann, Fink, Calzia, Burret, Radermacher, Tuckermann and Vettorazzi.)

Background: Sodium thiosulfate (STS) is a recognized drug with antioxidant and H 2 S releasing properties. We recently showed that STS attenuated organ dysfunction and injury during resuscitation from trauma-and-hemorrhage in CSE-ko mice, confirming its previously described organ-protective and anti-inflammatory properties. The role of H 2 S in diabetes mellitus type 1 (DMT1) is controversial: genetic DMT1 impairs H 2 S biosynthesis, which has been referred to contribute to endothelial dysfunction and cardiomyopathy. In contrast, development and severity of hyperglycemia in streptozotocin(STZ)-induced DMT1 was attenuated in CSE-ko mice. Therefore, we tested the hypothesis whether STS would also exert organ-protective effects in CSE-ko mice with STZ-induced DMT1, similar to our findings in animals without underlying co-morbidity., Methods: Under short-term anesthesia with sevoflurane and analgesia with buprenorphine CSE-ko mice underwent DMT1-induction by single STZ injection (100 μg⋅g -1 ). Seven days later, animals underwent blast wave-induced blunt chest trauma and surgical instrumentation followed by 1 h of hemorrhagic shock (MAP 35 ± 5 mmHg). Resuscitation comprised re-transfusion of shed blood, lung-protective mechanical ventilation, fluid resuscitation and continuous i.v. norepinephrine together with either i.v. STS (0.45 mg⋅g -1 ) or vehicle ( n = 9 in each group). Lung mechanics, hemodynamics, gas exchange, acid-base status, stable isotope-based metabolism, and visceral organ function were assessed. Blood and organs were collected for analysis of cytokines, chemokines, and immunoblotting., Results: Diabetes mellitus type 1 was associated with more severe circulatory shock when compared to our previous study using the same experimental design in CSE-ko mice without co-morbidity. STS did not exert any beneficial therapeutic effect. Most of the parameters measured of the inflammatory response nor the tissue expression of marker proteins of the stress response were affected either., Conclusion: In contrast to our previous findings in CSE-ko mice without underlying co-morbidity, STS did not exert any beneficial therapeutic effect in mice with STZ-induced DMT1, possibly due to DMT1-related more severe circulatory shock. This result highlights the translational importance of both integrating standard ICU procedures and investigating underlying co-morbidity in animal models of shock research., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Gröger, Hogg, Abdelsalam, Kress, Hoffmann, Stahl, Calzia, Wachter, Vogt, Wang, Merz, Radermacher and McCook.)

Objectives: Glucocorticoids (GCs) are one of the most widely prescribed anti-inflammatory drugs. By acting through their cognate receptor, the glucocorticoid receptor (GR), GCs downregulate the expression of pro-inflammatory genes and upregulate the expression of anti-inflammatory genes. Metabolic pathways have recently been identified as key parts of both the inflammatory activation and anti-inflammatory polarization of macrophages, immune cells responsible for acute inflammation and tissue repair. It is currently unknown whether GCs control macrophage metabolism, and if so, to what extent metabolic regulation by GCs confers anti-inflammatory activity., Methods: Using transcriptomic and metabolomic profiling of macrophages, we identified GC-controlled pathways involved in metabolism, especially in mitochondrial function., Results: Metabolic analyses revealed that GCs repress glycolysis in inflammatory myeloid cells and promote tricarboxylic acid (TCA) cycle flux, promoting succinate metabolism and preventing intracellular accumulation of succinate. Inhibition of ATP synthase attenuated GC-induced transcriptional changes, likely through stalling of TCA cycle anaplerosis. We further identified a glycolytic regulatory transcription factor, HIF1α, as regulated by GCs, and as a key regulator of GC responsiveness during inflammatory challenge., Conclusions: Our findings link metabolism to gene regulation by GCs in macrophages., (Copyright © 2021 The Author(s). Published by Elsevier GmbH.. All rights reserved.)

Background: Sodium thiosulfate (Na2S2O3) is a clinically established drug with antioxidant and sulphide-releasing properties. Na2S2O3 mediated neuro- and cardioprotective effects in ischemia/reperfusion models and anti-inflammatory effects in LPS-induced acute lung injury. Moreover, Na2S2O3 improved lung function during resuscitation from hemorrhagic shock in swine with pre-existing atherosclerosis, characterized by decreased expression of cystathionine γ-lyase (CSE), a major source of hydrogen sulfide (H2S) synthesis in the vasculature. Based on these findings, we investigated the effects of Na2S2O3 administration during resuscitation from trauma-and-hemorrhage in mice under conditions of whole body CSE deficit., Methods: After blast wave-induced blunt chest trauma and surgical instrumentation, CSE knockout (CSE-/-) mice underwent 1 h of hemorrhagic shock (MAP 35 ± 5 mm Hg). At the beginning of resuscitation comprising retransfusion, norepinephrine support and lung-protective mechanical ventilation, animals received either i.v. Na2S2O3 (0.45 mg g-1, n = 12) or vehicle (saline, n = 13). Hemodynamics, acid-base status, metabolism using stable isotopes, and visceral organ function were assessed. Blood and organs were collected for analysis of cytokines, mitochondrial respiratory capacity, and immunoblotting., Results: Na2S2O3 treatment improved arterial paO2 (P = 0.03) coinciding with higher lung tissue glucocorticoid receptor expression. Norepinephrine requirements were lower in the Na2S2O3 group (P < 0.05), which was associated with lower endogenous glucose production and higher urine output. Na2S2O3 significantly increased renal tissue IκBα and heme oxygenase-1 expression, whereas it lowered kidney IL-6 and MCP-1 levels., Conclusion: Na2S2O3 exerted beneficial effects during resuscitation of murine trauma-and-hemorrhage in CSE-/- mice, confirming and extending the previously described organ-protective and anti-inflammatory properties of Na2S2O3. The findings make Na2S2O3 a potentially promising therapeutic option in the context of impaired CSE activity and/or reduced endogenous H2S availability., Competing Interests: The authors report no conflicts of interest., (Copyright © 2021 by the Shock Society.)

Abdominal trauma (AT) is of major global importance, particularly with the increased potential for civil, terroristic, and military trauma. The injury pattern and systemic consequences of blunt abdominal injuries are highly variable and frequently underestimated or even missed, and the pathomechanisms remain still poorly understood. Therefore, we investigated the temporal-spatial organ and immune response after a standardized blast-induced blunt AT. Anesthetized mice were exposed to a single blast wave centered on the epigastrium. At 2, 6, or 24 h after trauma, abdominal organ damage was assessed macroscopically, microscopically, and biochemically. A higher degree of trauma severity, determined by a reduction of the distance between the epigastrium and blast inductor, was reflected by a reduced survival rate. The hemodynamic monitoring during the first 120 min after AT revealed a decline in the mean arterial pressure within the first 80 min, whereas the heart rate remained quite stable. AT induced a systemic damage and inflammatory response, evidenced by elevated HMGB-1 and IL-6 plasma levels. The macroscopic injury pattern of the abdominal organs (while complex) was consistent, with the following frequency: liver > pancreas > spleen > left kidney > intestine > right kidney > others > lungs and was reflected by microscopic liver and pancreas damages. Plasma levels of organ dysfunction markers increased during the first 6 h after AT and subsequently declined, indicating an early, temporal impairment of the function on a multi-organ level. The established highly reproducible murine blunt AT, with time- and trauma-severity-dependent organ injury patterns, systemic inflammatory response, and impairment of various organ functions, reflects characteristics of human AT. In the future, this model may help to study the complex immuno-pathophysiological consequences and innovative therapeutic approaches after blunt AT., (© 2021 The Authors. The FASEB Journal published by Wiley Periodicals LLC on behalf of Federation of American Societies for Experimental Biology.)

Severe injuries are frequently accompanied by hemorrhagic shock and harbor an increased risk for complications. Local or systemic inflammation after trauma/hemorrhage may lead to a leaky intestinal epithelial barrier and subsequent translocation of gut microbiota, potentially worsening outcomes. To evaluate the extent with which trauma affects the gut microbiota composition, we performed a post hoc analysis of a murine model of polytrauma and hemorrhage. Four hours after injury, organs and plasma samples were collected, and the diversity and composition of the cecal microbiome were evaluated using 16S rRNA gene sequencing. Although cecal microbial alpha diversity and microbial community composition were not found to be different between experimental groups, norepinephrine support in shock animals resulted in increased alpha diversity, as indicated by higher numbers of distinct microbial features. We observed that the concentrations of proinflammatory mediators in plasma and intestinal tissue were associated with measures of microbial alpha and beta diversity and the presence of specific microbial drivers of inflammation, suggesting that the composition of the gut microbiome at the time of trauma, or shortly after trauma exposure, may play an important role in determining physiological outcomes. In conclusion, we found associations between measures of gut microbial alpha and beta diversity and the severity of systemic and local gut inflammation. Furthermore, our data suggest that four hours following injury is too early for development of global changes in the alpha diversity or community composition of the intestinal microbiome. Future investigations with increased temporal-spatial resolution are needed in order to fully elucidate the effects of trauma and shock on the gut microbiome, biological signatures of inflammation, and proximal and distal outcomes.

Abstract: In activated immune cells, differentiation and function are determined by cell type-specific modifications of metabolic patterns. After traumatic brain injury both immune cell activation and suppression were reported. Therefore, we sought to explore immune cell energy metabolism in a long-term, resuscitated porcine model of acute subdural hematoma (ASDH)-induced acute brain injury devoid of impaired systemic hemodynamics and oxygen transport.Before and up to 50 h after induction of ASDH, peripheral blood mononuclear cells (PBMCs) were separated by density gradient centrifugation, and cell metabolism was analyzed using high-resolution respirometry for mitochondrial respiration and electron spin resonance for reactive oxygen species production. After incubation with stable isotope-labeled 1,2-13C2-glucose or 13C5-glutamine, distinct labeling patterns of intermediates of glycolysis or tricarboxylic acid (TCA) cycle and 13CO2 production were measured by gas chromatography-mass spectroscopy. Principal component analysis was followed by a varimax rotation on the covariance across all measured variables and all measured time points.After ASDH induction, average PBMC metabolic activity remained unaffected, possibly because strict adherence to intensive care unit guidelines limited trauma to ASDH induction without any change in parameters of systemic hemodynamics, oxygen transport, and whole-body metabolism. Despite decreased glycolytic activity fueling the TCA cycle, the principal component analysis indicated a cell type-specific activation pattern with biosynthetic and proliferative characteristics., Competing Interests: The authors report no conflicts of interest., (Copyright © 2020 by the Shock Society.)

In studies that target specific functions or organs, the response is often overlaid by indirect effects of the intervention on global metabolism. The metabolic side of these interactions can be assessed based on total energy expenditure (TEE) and the contributions of the principal energy sources, carbohydrates, proteins and fat to whole body CO 2 production. These parameters can be identified from indirect calorimetry using respiratory oxygen intake and CO 2 dioxide production data that are combined with the response of the 13 CO 2 release in the expired air and the glucose tracer enrichment in plasma following a 13 C glucose stable isotope infusion. This concept is applied to a mouse protocol involving anesthesia, mechanical respiration, a disease model, like hemorrhage and therapeutic intervention. It faces challenges caused by a small sample size for both breath and plasma as well as changes in metabolic parameters caused by disease and intervention. Key parameters are derived from multiple measurements, all afflicted with errors that may accumulate leading to unrealistic values. To cope with these challenges, a sensitive on-line breath analysis system based on substrate-integrated hollow waveguide infrared spectroscopy and luminescence (iHWG-IR-LS) was used to monitor gas exchange values. A Bayesian statistical model is developed that uses established equations for indirect calorimetry to predict values for respiratory gas exchange and tracer data that are consistent with the corresponding measurements and also provides statistical error bands for these parameters. With this new methodology, it was possible to estimate important metabolic parameters (respiratory quotient (RQ), relative contribution of carbohydrate, protein and fat oxidation f carb , f fat and f prot , total energy expenditure TEE) in a resolution never available before for a minimal invasive protocol of mice under anesthesia.

Competing Interests: The authors report no conflicts of interest.

Immune cell activation leads to the acquisition of new functions, such as proliferation, chemotaxis, and cytokine production. These functional changes require continuous metabolic adaption in order to sustain ATP homeostasis for sufficient host defense. The bioenergetic demands are usually met by the interconnected metabolic pathways glycolysis, TCA cycle, and oxidative phosphorylation. Apart from glucose, other sources, such as fatty acids and glutamine, are able to fuel the TCA cycle.Rising evidence has shown that cellular metabolism has a direct effect on the regulation of immune cell functions. Thus, quiescent immune cells maintain a basal metabolic state, which shifts to an accelerated metabolic level upon immune cell activation in order to promote key effector functions.This review article summarizes distinct metabolic signatures of key immune cell subsets from quiescence to activation and demonstrates a methodical concept of how to assess cellular metabolic pathways. It further discusses why metabolic functions are of rising interest for translational research and how they can be affected by the underlying pathophysiological condition and/or therapeutic interventions.

Trauma represents a major socioeconomic burden worldwide. After a severe injury, hemorrhagic shock (HS) as a frequent concomitant aspect is a central driver of systemic inflammation and organ damage. The kidney is often strongly affected by traumatic-HS, and acute kidney injury (AKI) poses the patient at great risk for adverse outcome. Recently, thirty-eight-negative kinase 1 (TNK1) was proposed to play a detrimental role in organ damage after trauma/HS. Therefore, we aimed to assess the role of TNK1 in HS-induced kidney injury in a murine and a post hoc analysis of a non-human primate model of HS comparable to the clinical situation. Mice and non-human primates underwent resuscitated HS at 30 mmHg for 60 min. 5 h after the induction of shock, animals were assessed for systemic inflammation and TNK1 expression in the kidney. In vitro , murine distal convoluted tubule cells were stimulated with inflammatory mediators to gain mechanistic insights into the role of TNK1 in kidney dysfunction. In a translational approach, we investigated blood drawn from either healthy volunteers or severely injured patients at different time points after trauma (from arrival at the emergency room and at fixed time intervals until 10 days post injury; identifier: NCT02682550, https://clinicaltrials.gov/ct2/show/NCT02682550). A pronounced inflammatory response, as seen by increased IL-6 plasma levels as well as early signs of AKI, were observed in mice, non-human primates, and humans after trauma/HS. TNK1 was found in the plasma early after trauma-HS in trauma patients. Renal TNK1 expression was significantly increased in mice and non-human primates after HS, and these effects with concomitant induction of apoptosis were blocked by therapeutic inhibition of complement C3 activation in non-human primates. Mechanistically, in vitro data suggested that IL-6 rather than C3 cleavage products induced upregulation of TNK1 and impaired barrier function in renal epithelial cells. In conclusion, these data indicate that C3 inhibition in vivo may inhibit an excessive inflammatory response and mediator release, thereby indirectly neutralizing TNK1 as a potent driver of organ damage. In future studies, we will address the therapeutic potential of direct TNK1 inhibition in the context of severe tissue trauma with different degrees of additional HS., (Copyright © 2020 Halbgebauer, Karasu, Braun, Palmer, Braumüller, Schultze, Schäfer, Bückle, Eigner, Wachter, Radermacher, Resuello, Tuplano, Nilsson Ekdahl, Nilsson, Armacki, Kleger, Seufferlein, Kalbitz, Gebhard, Lambris, van Griensven and Huber-Lang.)

The metabolism can be explored via 13 C labeling of biological active substances and subsequent quantification of 13 C enrichment in the exhaled carbon dioxide in breath. The resulting tracer enrichment values can be determined by Fourier-transform Infrared Spectroscopy (FTIR), since different CO 2 isotopologues result in distinct absorption lines in the spectrum.The corresponding determination poses two challenges: first, FTIR absorbance can contain a nonlinear relationship between analyte amount and spectral signal and second, the spectral peaks for the different isotopologues overlap. The overlap precludes a separate calibration to asses the isotopologue concentration values and with it a determination of enrichments from concentration values. We propose here, first, a data reduction step like Principal Component Analysis (PCA) to convert the spectral information into one score pertaining to the 13 CO 2 enrichment. In a second step, a calibration function between score and enrichment values was established. The enrichment score can be derived by normalizing a subset of the spectrum by some measure for the 12 CO 2 sample content. Alternatively, the overlapping spectra were decomposed into two isotopologue spectra and the intensity of the separated spectra was used to form an enrichment score. For spectral separation, either Multivariate Curve Resolution Alternating Least Squares (MCR-ALS) was used or a novel decomposition strategy developed for this paper called Rotation and Angle-Bending Bayesian induced Transformation - Multivariate Curve Resolution (RABBIT - MCR) that operates in a Principal Component Analysis (PCA) subspace and is derived from MCR. We compared 13 C enrichment estimates from FTIR CO 2 spectra using different normalization variants with the two spectral separation models. In conclusion, the two spectral separation variants performed nearly equal, but better than any normalization variant., (Copyright © 2019. Published by Elsevier B.V.)

Background: Sepsis, that can be modeled by LPS injections, as an acute systemic inflammation syndrome is the most common cause for acute lung injury (ALI). ALI induces acute respiratory failure leading to hypoxemia, which is often associated with multiple organ failure (MOF). During systemic inflammation, the hypothalamus-pituitary-adrenal axis (HPA) is activated and anti-inflammatory acting glucocorticoids (GCs) are released to overcome the inflammation. GCs activate the GC receptor (GR), which mediates its effects via a GR monomer or GR dimer. The detailed molecular mechanism of the GR in different inflammatory models and target genes that might be crucial for resolving inflammation is not completely identified. We previously observed that mice with attenuated GR dimerization (GR dim/dim ) had a higher mortality in a non-resuscitated lipopolysaccharide (LPS)- and cecal ligation and puncture (CLP)-induced inflammation model and are refractory to exogenous GCs to ameliorate ALI during inflammation. Therefore, we hypothesized that impaired murine GR dimerization (GR dim/dim ) would further impair organ function in LPS-induced systemic inflammation under human like intensive care management and investigated genes that are crucial for lung function in this setup. Methods: Anesthetized GR dim/dim and wildtype (GR +/+ ) mice were challenged with LPS (10 mg·kg -1 , intraperitoneal) and underwent intensive care management ("lung-protective" mechanical ventilation, crystalloids, and norepinephrine) for 6 h. Lung mechanics and gas exchange were assessed together with systemic hemodynamics, acid-base status, and mitochondrial oxygen consumption (JO 2 ). Western blots, immunohistochemistry, and real time quantitative polymerase chain reaction were performed to analyze lung tissue and inflammatory mediators were analyzed in plasma and lung tissue. Results: When animals were challenged with LPS and subsequently resuscitated under intensive care treatment, GR dim/dim mice had a higher mortality compared to GR +/+ mice, induced by an increased need of norepinephrine to achieve hemodynamic targets. After challenge with LPS, GR dim/dim mice also displayed an aggravated ALI shown by a more pronounced impairment of gas exchange, lung mechanics and increased osteopontin (Opn) expression in lung tissue. Conclusion: Impairment of GR dimerization aggravates systemic hypotension and impairs lung function during LPS-induced endotoxic shock in mice. We demonstrate that the GR dimer is an important mediator of hemodynamic stability and lung function, possibly through regulation of Opn, during LPS-induced systemic inflammation., (Copyright © 2020 Wepler, Preuss, Merz, Hartmann, Wachter, McCook, Vogt, Kress, Gröger, Fink, Scheuerle, Möller, Calzia, Burret, Radermacher, Tuckermann and Vettorazzi.)

Controversial data are available on hydrogen sulfide (H 2 S) during hemorrhage and resuscitation, depending on timing, dosing, mode of application, and the H 2 S donor used. Sodium thiosulfate (Na 2 S 2 O 3 ) is a recognized drug devoid of major side effects, which attenuated murine acute lung injury and cerebral ischemia/reperfusion injury. Therefore, we tested the hypothesis whether Na 2 S 2 O 3 would mitigate organ dysfunction in porcine hemorrhage-and-resuscitation. We studied animals with pre-existing coronary artery disease because of the reduced coronary arterial expression of the H 2 S producing enzyme cystathionine-γ-lyase (CSE) in this prospective, randomized, controlled, blinded experimental study. 20 anesthetized and instrumented pigs underwent 3 h of hemorrhage (removal of 30 % of the blood volume and subsequent titration of mean arterial pressure to 40 mmHg). Resuscitation (72 h) comprised re-transfusion of shed blood, crystalloids, and continuous i.v. norepinephrine. Animals randomly received vehicle or Na 2 S 2 O 3 (0.1 g·kg -1  h -1 ) for 24 h. Before, at the end of and every 24 h after shock, hemodynamics, metabolism, blood gases, lung, heart, kidney, and liver function and injury were evaluated together with cytokines and parameters of oxidative and nitrosative stress. Immediate post mortem lung, kidney, heart, and liver specimen were analyzed for marker proteins of inflammation and oxidative and nitrosative stress and mitochondrial respiratory activity in the heart, kidney, and liver. Immuno-histochemical analysis comprised lung extra-vascular albumin accumulation, nitrotyrosine formation, and CSE and glucocorticoid receptor (GCR) expression. Na 2 S 2 O 3 significantly attenuated shock-induced impairment of lung mechanics and gas exchange (plateau and positive end-expiratory pressure at 72 h p = 0.0006/p = 0.0264; Horovitz index at 48 h p = 0.0261), which coincided with a higher tissue GCR expression (p = 0.0415). During resuscitation from hemorrhagic shock Na 2 S 2 O 3 attenuated shock-induced acute lung injury in co-morbid swine, most likely due to a GCR expression related mechanism., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Hyperoxia (ventilation with FIO2 = 1.0) has vasoconstrictor properties, in particular in the coronary vascular bed, and, hence, may promote cardiac dysfunction. However, we previously showed that hyperoxia attenuated myocardial injury during resuscitation from hemorrhage in swine with coronary artery disease. Therefore, we tested the hypothesis whether hyperoxia would also mitigate myocardial injury and improve heart function in the absence of chronic cardiovascular comorbidity.After 3 h of hemorrhage (removal of 30% of the calculated blood volume and subsequent titration of mean arterial pressure to 40 mm Hg) 19 anesthetized, mechanically ventilated, and instrumented pigs received FIO2 = 0.3(control) or hyperoxia(FIO2 = 1.0) during the first 24 h. Before, at the end of and every 12 h after shock, hemodynamics, blood gases, metabolism, cytokines, and cardiac function (pulmonary artery thermodilution, left ventricular pressure-conductance catheterization) were recorded. At 48 h, cardiac tissue was harvested for western blotting, immunohistochemistry, and mitochondrial respiration.Except for higher left ventricular end-diastolic pressures at 24 h (hyperoxia 21 (17;24), control 17 (15;18) mm Hg; P = 0.046), hyperoxia affected neither left ventricular function cardiac injury (max. Troponin I at 12 h: hyperoxia:9 (6;23), control:17 (11;24) ng mL; P = 0.395), nor plasma cytokines (except for interleukin-1β: hyperoxia 10 (10;10) and 10 (10;10)/control 14 (10;22), 12 (10;15) pg mL, P = 0.023 and 0.021 at 12 and 24 h, respectively), oxidation and nitrosative stress, and mitochondrial respiration. However, hyperoxia decreased cardiac tissue three-nitrotyrosine formation (P < 0.001) and inducible nitric oxide synthase expression (P = 0.016). Ultimately, survival did not differ significantly either.In conclusion, in contrast to our previous study in swine with coronary artery disease, hyperoxia did not beneficially affect cardiac function or tissue injury in healthy swine, but was devoid of deleterious side effects.

Hemorrhagic shock (HS) accounts for 30% to 40% of trauma-induced mortality, which is due to multi-organ-failure subsequent to systemic hyper-inflammation, triggered by hypoxemia and tissue ischemia. The slow-releasing, mitochondria-targeted H2S donor AP39 exerted beneficial effects in several models of ischemia-reperfusion injury and acute inflammation. Therefore, we tested the effects of AP39-treatment in a murine model of combined blunt chest trauma (TxT) and HS with subsequent resuscitation., Methods: After blast wave-induced TxT or sham procedure, anesthetized and instrumented mice underwent 1 h of hemorrhage followed by 4 h of resuscitation comprising an i.v. bolus injection of 100 or 10 nmol kg AP39 or vehicle, retransfusion of shed blood, fluid resuscitation, and norepinephrine. Lung mechanics and gas exchange were assessed together with hemodynamics, metabolism, and acid-base status. Blood and tissue samples were analyzed for cytokine and chemokine levels, western blot, immunohistochemistry, mitochondrial oxygen consumption (JO2), and histological changes., Results: High dose AP39 attenuated systemic inflammation and reduced the expression of inducible nitric oxide synthase (iNOS) and IκBα expression in lung tissue. In the combined trauma group (TxT + HS), animals treated with high dose AP39 presented with the lowest mean arterial pressure and thus highest norepinephrine requirements and higher mortality. Low dose AP39 had no effects on hemodynamics, leading to unchanged norepinephrine requirements and mortality rates., Conclusion: AP39 is a systemic anti-inflammatory agent. In our model of trauma with HS, there may be a narrow dosing and timing window due to its potent vasodilatory properties, which might result in or contribute to aggravation of circulatory shock-related hypotension.

Background: Hypoxemia and tissue ischemia during hemorrhage as well as formation of oxygen and nitrogen radicals during resuscitation promote hyperinflammation and, consequently, trigger severe multi-organ failure (MOF). Individuals diagnosed with stress-related disorders or reporting a life history of psychosocial stress are characterized by chronic low-grade inflammation and a reduced glucocorticoid (GC) signaling. We hypothesized that exposure to chronic psychosocial stress during adulthood prior to hemorrhagic shock increases oxidative/nitrosative stress and therefore the risk of developing MOF in mice., Methods and Findings: To induce chronic psychosocial stress linked to mild immune activation and reduced GC signaling in male mice, the chronic subordinate colony housing (CSC) paradigm was employed. Single-housed (SHC) mice were used as controls. Subsequently, CSC and SHC mice were exposed to hemorrhagic shock following resuscitation to investigate the effects of prior psychosocial stress load on survival, organ function, metabolism, oxidative/nitrosative stress, and inflammatory readouts. An increased adrenal weight in CSC mice indicates that the stress paradigm reliably worked. However, no effect of prior psychosocial stress on outcome after subsequent hemorrhage and resuscitation could be detected., Conclusions: Chronic psychosocial stress during adulthood is not sufficient to promote hemodynamic complications, organ dysfunction, metabolic disturbances and did not increase the risk of MOF after subsequent hemorrhage and resuscitation. Intravenous norepinephrine to keep target hemodynamics might have led to a certain level of oxidative stress in both groups and, therefore, disguised potential effects of chronic psychosocial stress on organ function after hemorrhagic shock in the present murine trauma model.

Introduction: Hemorrhagic shock is a major cause of death after trauma. An additional blunt chest trauma independently contributes to mortality upon the development of an acute lung injury (ALI) by aggravating pathophysiological consequences of hemorrhagic shock. The maintenance of hydrogen sulfide availability is known to play an important role during hemorrhage and ALI. We therefore tested the impact of a genetic 3-mercaptopyruvate sulfurtransferase mutation (Δ3-MST) in a resuscitated murine model of traumatic-hemorrhagic shock., Methods: Anesthetized wild-type (WT) and Δ3-MST mice underwent hemorrhagic shock with/without blunt chest trauma. Hemorrhagic shock was implemented for 1 h followed by retransfusion of shed blood and intensive care therapy for 4 h, including lung-protective mechanical ventilation, fluid resuscitation, and noradrenaline titrated to maintain a mean arterial pressure at least 50 mmHg. Systemic hemodynamics, metabolism, and acid-base status were assessed together with lung mechanics and gas exchange. Postmortem tissue samples were analyzed for immunohistological protein expression and mitochondrial oxygen consumption., Results: 3-MST-deficient mice showed similar results in parameters of hemodynamics, gas exchange, metabolism, acid base status, and survival compared with the respective WT controls. Renal albumin extravasation was increased in Δ3-MST mice during hemorrhagic shock, together with a decrease of LEAK respiration in heart tissue. In contrast, mitochondrial oxygen consumption in the uncoupled state was increased in kidney and liver tissue of Δ3-MST mice subjected to the combined trauma., Conclusions: In summary, in a resuscitated murine model of traumatic-hemorrhagic shock, 3-MST deficiency had no physiologically relevant impact on hemodynamics and metabolism, which ultimately lead to unchanged mortality regardless of an additional blunt chest trauma.

Introduction: Hemorrhagic shock accounts for a large amount of trauma-related mortality. The severity of trauma can be further aggravated by an additional blunt chest trauma (TxT), which independently contributes to mortality upon the development of an acute lung injury (ALI). Besides, cigarette smoke (CS) exposure before TxT enhanced posttraumatic inflammation, thereby aggravating ALI. We therefore aimed to characterize the impact of an acute and/or chronic lung injury on organ dysfunction in a murine model of traumatic hemorrhagic shock (HS)., Methods: After 3 weeks of CS exposure, anesthetized mice underwent HS with/without TxT. Hemorrhagic shock was implemented for 1 h followed by retransfusion of shed blood and intensive care therapy for 4 h including lung-protective mechanical ventilation, fluid resuscitation, and noradrenaline titrated to maintain mean arterial pressure ≥50 mmHg. Lung mechanics and gas exchange were assessed together with systemic hemodynamics, metabolism, and acid-base status. Postmortem blood and tissue samples were analyzed for cytokine and chemokine levels, protein expression, mitochondrial respiration, and histological changes., Results: CS exposure and HS alone coincided with increased inflammation, decreased whole blood sulfide concentrations, and decreased diaphragmatic mitochondrial respiration. CS-exposed mice, which were subjected to TxT and subsequent HS, showed hemodynamic instability, acute kidney injury, and high mortality., Conclusions: Chronic CS exposure per se had the strongest impact on inflammatory responses. The degree of inflammation was similar upon an additional TxT, however, mice presented with organ dysfunction and increased mortality rates. Hence, in mice the degree of inflammation may be dissociated from the severity of organ dysfunction or injury.

A multiparameter gas sensor based on distributed feedback interband cascade lasers emitting at 4.35 μm and ultrafast electro-spun luminescence oxygen sensors has been developed for the quantification and continuous monitoring of 13 CO 2 / 12 CO 2 isotopic ratio changes and oxygen in exhaled mouse breath samples. Mid-infrared absorption spectra for quantitatively monitoring the enrichment of 13 CO 2 levels were recorded in a miniaturized dual-channel substrate-integrated hollow waveguide using balanced ratiometric detection, whereas luminescence quenching was used for synchronously detecting exhaled oxygen levels. Allan variance analysis verified a CO 2 measurement precision of 1.6‰ during a 480 s integration time. Routine online monitoring of exhaled mouse breath was performed in 14 mechanically ventilated and instrumented mice and demonstrated the feasibility of online isotope-selective exhaled breath analysis within microliters of probed gas samples using the reported combined sensor platform.

Exhaled breath offers monitoring bio markers, as well as diagnosing diseases and therapeutic interventions. In addition, vital functions may be non-invasively monitored online. Animal models are frequently used in research for determining novel therapeutic approaches and/or for investigating biological pathways. The exhaled carbon dioxide concentration, exhaled and inhaled oxygen concentration, and the subsequent respiratory quotient (RQ) offer insight into metabolic activity. One may adapt breath sampling systems and equipment designed for human applications to large animal studies. However, such adaptations are usually impossible for small animals due to their minuscule breath volume. Here, we present a system for the online monitoring of exhaled breath in a 'mouse intensive care unit' (MICU) based on a modified Fourier-transform infrared spectrometer equipped with a substrate-integrated hollow waveguide gas cell, and a luminescence-based oxygen flow-through sensor integrated into the respiratory equipment of the MICU. Thereby, per-minute resolution of O 2 consumption and CO 2 production was obtained, and the 95% confidence range of the determined RQ was ±0.04 or approximately ±5% of the nominal value. Changes in the RQ value caused by intervention in either the metabolic or respiratory system may therefore reliably be detected.

Hemorrhagic shock (HS) after tissue trauma increases the complication and mortality rate of polytrauma (PT) patients. Although several murine trauma models have been introduced, there is a lack of knowledge about the exact impact of an additional HS. We hypothesized that HS significantly contributes to organ injury, which can be reliably monitored by detection of specific organ damage markers. Therefore we established a novel clinically relevant PT plus HS model in C57BL/6 mice which were randomly assigned to control, HS, PT, or PT+HS procedure (n = 8 per group). For induction of PT, anesthetized animals received a blunt chest trauma, head injury, femur fracture, and soft tissue injury. HS was induced by pressure-controlled blood drawing (mean arterial blood pressure of 30 mmHg for 60 min) and mice then resuscitated with ionosterile (4 × volume drawn), monitored, and killed for blood and organ harvesting 4 h after injury. After HS and resuscitation, PT+HS mice required earlier and overall more catecholamine support than HS animals to keep their mean arterial blood pressure. HS significantly contributed to the systemic release of interleukin-6 and high mobility group box 1 protein. Furthermore, the histological lung injury score, pulmonary edema, neutrophil influx, and plasma clara cell protein 16 were all significantly enhanced in PT animals in the presence of an additional HS. Although early morphological changes were minor, HS also contributed functionally to remote acute kidney injury but not to early liver damage. Moreover, PT-induced systemic endothelial injury, as determined by plasma syndecan-1 levels, was significantly aggravated by an additional HS. These results indicate that HS adds to the systemic inflammatory reaction early after PT. Within hours after PT, HS seems to aggravate pulmonary damage and to worsen renal and endothelial function which might overall contribute to the development of early multiple organ dysfunction.

Objectives: Investigation of the effects of hyperoxia during resuscitation from hemorrhagic shock in swine with preexisting coronary artery disease., Design: Prospective, controlled, randomized trial., Setting: University animal research laboratory., Subjects: Nineteen hypercholesterolemic pigs with preexisting coronary artery disease., Interventions: Anesthetized, mechanically ventilated, and surgically instrumented pigs underwent 3 hours of hemorrhagic shock (removal of 30% of the calculated blood volume and subsequent titration of mean arterial blood pressure ≈40 mm Hg). Postshock resuscitation (48 hr) comprised retransfusion of shed blood, crystalloids (balanced electrolyte solution), and norepinephrine support. Pigs were randomly assigned to "control" (FIO2 0.3, adjusted for arterial oxygen saturation ≥ 90%) and "hyperoxia" (FIO2 1.0 for 24 hr) groups., Measurements and Main Results: Before, at the end of shock and every 12 hours of resuscitation, datasets comprising hemodynamics, calorimetry, blood gases, cytokines, and cardiac and renal function were recorded. Postmortem, organs were sampled for immunohistochemistry, western blotting, and mitochondrial high-resolution respirometry. Survival rates were 50% and 89% in the control and hyperoxia groups, respectively (p = 0.077). Apart from higher relaxation constant τ at 24 hours, hyperoxia did not affect cardiac function. However, troponin values were lower (2.2 [0.9-6.2] vs 6.9 [4.8-9.8] ng/mL; p < 0.05) at the end of the experiment. Furthermore, hyperoxia decreased cardiac 3-nitrotyrosine formation and increased inducible nitric oxide synthase expression. Plasma creatinine values were lower in the hyperoxia group during resuscitation coinciding with significantly improved renal mitochondrial respiratory capacity and lower 3-nitrotyrosine formation., Conclusions: Hyperoxia during resuscitation from hemorrhagic shock in swine with preexisting coronary artery disease reduced renal dysfunction and cardiac injury, potentially resulting in improved survival, most likely due to increased mitochondrial respiratory capacity and decreased oxidative and nitrosative stress. Compared with our previous study, the present results suggest a higher benefit of hyperoxia in comorbid swine due to an increased susceptibility to hemorrhagic shock.

Background: Cystathionine-γ-lyase (CSE) was shown to have a regulatory role in glucose metabolism. Circulatory shock can induce metabolic stress, thereby leading to hyperglycemia and mitochondrial dysfunction. In vitro data suggest an effect of high glucose on CSE expression. Therefore, the aim of this study was to investigate the effects of hyperglycemia on CSE expression in resuscitated murine septic shock., Methods: Normo- (80-150 mg/dl) and hyperglycemic (>200 mg/dl) male C57/BL6J mice (n = 5-6 per group) underwent cecal ligation and puncture (CLP)-induced polymicrobial sepsis or sham procedure (n = 6 per group) and, 15 h afterwards, were anesthetized again, surgically instrumented and received intensive care treatment, including antibiotics, lung protective mechanical ventilation, circulatory support, and intravenous (i.v.) glucose infusion (50% as stable-isotope labeled 1,2,3,4,5,6- 13 C 6 glucose). Blood and breath gas were sampled hourly to quantify parameters of glucose metabolism. 5 h later, mice were sacrificed and organs were harvested. The liver mitochondrial respiratory activity was determined via high resolution respirometry; CSE, peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC1α), and adipocyte differentiation-related protein (ADRP) expression was immunohistochemically investigated., Results: In sepsis combined with hyperglycemia the least CSE and PGC1α expression could be detected, along with reduced mitochondrial respiratory activity, and enhanced ADRP expression, a marker of lipid droplet formation, in the liver. A novel in vivo finding is the CSE translocation from the cytosol to the nucleus triggered by metabolic stress., Conclusions: A relationship between CSE and glucose metabolism was established, which, when dysregulated, may contribute to fatty liver disease and hepatic steatosis.

We previously demonstrated beneficial effects of 22 h of hyperoxia following near-lethal porcine hemorrhagic shock, whereas therapeutic hypothermia was detrimental. Therefore, we investigated whether shorter exposure to hyperoxia (12 h) would still improve organ function, and whether 12 h of hypothermia with subsequent rewarming could avoid deleterious effects after less severe hemorrhagic shock.Twenty-seven anesthetized and surgically instrumented pigs underwent 3 h of hemorrhagic shock by removal of 30% of the blood volume and titration of the mean arterial blood pressure (MAP) to 40 mm Hg. Post-shock, pigs were randomly assigned to control, hyperoxia (FIO2 100% for 12 h) or hypothermia group (34°C core temperature for 12 h with subsequent rewarming). Before, at the end of shock, after 12 and 23 h of resuscitation, data sets comprising hemodynamics, blood gases, and parameters of inflammation and organ function were acquired. Postmortem, kidney samples were collected for immunohistochemistry and western blotting.Hyperoxia exerted neither beneficial nor detrimental effects. In contrast, mortality in the hypothermia group was significantly higher compared with controls (67% vs. 11%). Hypothermia impaired circulation (MAP 64 (57;89) mm Hg vs. 104 (98; 114) mm Hg) resulting in metabolic acidosis (lactate 11.0 (6.6;13.6) mmol L vs. 1.0 (0.8;1.5) mmol L) and reduced creatinine clearance (26 (9;61) mL min vs. 77 (52;80) mL min) compared to the control group after 12 h of resuscitation. Impaired kidney function coincided with increased renal 3-nitrotyrosine formation and extravascular albumin accumulation.In conclusion, hyperoxia proved to be safe during resuscitation from hemorrhagic shock. The lacking organ-protective effects of hyperoxia compared to resuscitation from near-lethal hemorrhage suggest a dependence of the effectiveness of hyperoxia from shock severity. In line with our previous report, therapeutic hypothermia (and rewarming) was confirmed to be detrimental most likely due to vascular barrier dysfunction.

Decreased levels of endogenous hydrogen sulfide (H2S) contribute to atherosclerosis, whereas equivocal data are available on H2S effects during sepsis. Moreover, H2S improved glucose utilization in anaesthetized, ventilated, hypothermic mice, but normothermia and/or sepsis blunted this effect. The metabolic effects of H2S in large animals are controversial. Therefore, we investigated the effects of the H2S donor GYY4137 during resuscitated, fecal peritonitis-induced septic shock in swine with genetically and diet-induced coronary artery disease (CAD). Twelve and 18 h after peritonitis induction, pigs received either GYY4137 (10 mg kg, n = 9) or vehicle (n = 8). Before, at 12 and 24 h of sepsis, we assessed left ventricular (pressure-conductance catheters) and renal (creatinine clearance, blood NGAL levels) function. Endogenous glucose production and glucose oxidation were derived from the plasma glucose isotope and the expiratory CO2/CO2 enrichment during continuous i.v. 1,2,3,4,5,6-C6-glucose infusion. GYY4137 significantly increased aerobic glucose oxidation, which coincided with higher requirements of exogenous glucose to maintain normoglycemia, as well as significantly lower arterial pH and decreased base excess. Apart from significantly lower cardiac eNOS expression and higher troponin levels, GYY4137 did not significantly influence cardiac and kidney function or the systemic inflammatory response. During resuscitated septic shock in swine with CAD, GYY4137 shifted metabolism to preferential carbohydrate utilization. Increased troponin levels are possibly due to reduced local NO availability. Cautious dosing, the timing of GYY4137 administration, and interspecies differences most likely account for the absence of any previously described anti-inflammatory or organ-protective effects of GYY4137 in this model.

Gastric emptying can be assessed by an oral administration of a 13 C labeled substrate and its response in the expiratory release of the oxidation product [Formula: see text]. Impaired gut function, reflected, for example, in an intolerance against enteral nutrition may delay or discontinue gastric emptying, potentially leading to multiple peaks in the time profile of expiration. The resulting profile cannot be analyzed by the usual data evaluation that is based on a 'beta exponential' (BEX) function. We developed a new approach that better reflects the underlying physiology. It allows a flexible time profile of gastric release and considers a transient [Formula: see text] retention in different compartments as well as an incomplete recovery of [Formula: see text] in the expiration. Parameters that describe the distribution/retention kinetics cannot be determined based on the same breath data that were used to estimate emptying. To enable the determination of the kinetic parameters, they were constrained to match published data using a Bayesian statistical analysis. The applicability of the new model was compared with BEX for healthy subjects. BEX fails to explain the observed data and, compared to the new approach, overestimates the speed of emptying. Predictive accuracy under impaired gastric motility was explored using synthetic data. Only the new approach can reproduce a multiphase absorption profile. When routine benchtop equipment was used for measurements, then the rate-limiting step for precision in the estimate of emptying is the quality in the a priori estimate for kinetic parameters rather than precision in measurements. Only about 80% of the absorbed [Formula: see text] has to be released by expiration. With these features, the new approach promises to widen the applicability of breath tests for gastric emptying.

Backgrounds and Objectives: As part of the expansion of the site-specific education profile of the medical curriculum MED@ULM of the University of Ulm, a new track "trauma care and trauma research" was established in the winter semester 2012/2013. The acceptance of the track was evaluated during the winter semester 2013/2014., Material and Methods: The 6-semester track extends the existing curriculum by offering subjects in trauma management and trauma research to students of human medicine. A central aim of the track is to promote medical professional competence, expertise in emergency care and competence in trauma-related scientific work and research. Central learning contents could be intensified in newly established emergency simulation training. Additionally, participating students have to perform a doctoral thesis on an obligatory trauma-related experimental subject. A first analysis study focusing on the learning style of the participating students (n = 17) and a control group consisting of members of the same semester (n = 20) was performed using the Kolb learning style inventory. In a validated evaluation in the winter semesters 2013/2014 and 2014/2015, the students were asked about their expectations and experience with the track, criticisms, suggestions and satisfaction with the study conditions. The data were analyzed using descriptive statistics., Results: The analysis of the students' preferred learning styles revealed no differences between track students and the control group. Most of the students considered the track as a form of personal further education. The students had high expectations of practical skills with relevance to the clinical daily routine, learning scientific methods and preparing their thesis. The track students were more critical with regard to the study conditions than the control group students, although the track students of the third semester still judged their studies to be more interesting than the track students of the first semester and the control group., Conclusion: With the introduction of the new trauma track into the curriculum of the medical curriculum MED@ULM of the University of Ulm, a further possibility for medical students to focus on their own individual options was established. At least half of the track students wanted to be later active in the triad of patient care, teaching and research. Further investigations are necessary to determine whether the establishment of the trauma track has a positive influence on the number of new recruits in trauma surgery and anesthesiology.

Septic shock-related kidney failure is characterized by almost normal morphological appearance upon pathological examination. Endothelial barrier disrupture has been suggested to be of crucial importance for septic shock-induced organ dysfunction. Therefore, in murine resuscitated cecal ligation and puncture (CLP)-induced septic shock, we tested the hypothesis whether there is a direct relationship between the kidney endothelial barrier injury and renal dysfunction. Anesthetized mice underwent CLP, and 15 h later, were anesthetized again and surgically instrumented for a 5-h period of intensive care comprising lung-protective mechanical ventilation, fluid resuscitation, continuous i.v. norepinephrine to maintain target hemodynamics, and measurement of creatinine clearance (CrCl). Animals were stratified according to low or high CrCl. Nitrotyrosine formation, expression of the inducible isoform of the nitric oxide synthase, and blood cytokine (tumor necrosis factor, interleukin-6, interleukin-10) and chemokine (monocyte chemoattractant protein-1, keratinocyte-derived chemokine) levels were significantly higher in animals with low CrCl. When plotted against CrCl and neutrophil gelatinase-associated lipocalin levels, extravascular albumin accumulation, and tissue expression of the vascular endothelial growth factor and angiopoietin-1 showed significant mathematical relationships related to kidney (dys)function. Preservation of the constitutive expression of the hydrogen sulfide producing enzyme cystathione-γ-lyase was associated with maintenance of organ function. The direct quantitative relation between microvascular leakage and kidney (dys)function may provide a missing link between near-normal tissue morphology and septic shock-related renal failure, thus further highlighting the important role of vascular integrity in septic shock-related renal failure.

Hydrogen sulfide (H2S) is a gaseous signalling molecule involved in many physiological and pathological processes. There is increasing evidence that H2S is implicated in aging and lifespan control in the diet-induced longevity models. However, blood sulfide concentration of naturally long-lived species is not known. Here we measured blood sulfide in the long-lived naked mole-rat and five other mammalian species considerably differing in lifespan and found a negative correlation between blood sulfide and maximum longevity residual. In addition, we show that the naked mole-rat cystathionine β-synthase (CBS), an enzyme whose activity in the liver significantly contributes to systemic sulfide levels, has lower activity in the liver and is activated to a higher degree by S-adenosylmethionine compared to other species. These results add complexity to the understanding of the role of H2S in aging and call for detailed research on naked mole-rat transsulfuration., (Copyright © 2016 The Authors. Published by Elsevier B.V. All rights reserved.)

Objective: Hemorrhagic shock-induced tissue hypoxia induces hyperinflammation, ultimately causing multiple organ failure. Hyperoxia and hypothermia can attenuate tissue hypoxia due to increased oxygen supply and decreased demand, respectively. Therefore, we tested the hypothesis whether mild therapeutic hypothermia and hyperoxia would attenuate postshock hyperinflammation and thereby organ dysfunction., Design: Prospective, controlled, randomized study., Setting: University animal research laboratory., Subjects: Thirty-six Bretoncelles-Meishan-Willebrand pigs of either gender., Interventions: After 4 hours of hemorrhagic shock (removal of 30% of the blood volume, subsequent titration of mean arterial pressure at 35 mm Hg), anesthetized and instrumented pigs were randomly assigned to "control" (standard resuscitation: retransfusion of shed blood, fluid resuscitation, norepinephrine titrated to maintain mean arterial pressure at preshock values, mechanical ventilation titrated to maintain arterial oxygen saturation > 90%), "hyperoxia" (standard resuscitation, but FIO2, 1.0), "hypothermia" (standard resuscitation, but core temperature 34°C), or "combi" (hyperoxia plus hypothermia) (n = 9 each)., Measurements and Main Results: Before, immediately at the end of and 12 and 22 hours after hemorrhagic shock, we measured hemodynamics, blood gases, acid-base status, metabolism, organ function, cytokine production, and coagulation. Postmortem kidney specimen were taken for histological evaluation, immunohistochemistry (nitrotyrosine, cystathionine γ-lyase, activated caspase-3, and extravascular albumin), and immunoblotting (nuclear factor-κB, hypoxia-inducible factor-1α, heme oxygenase-1, inducible nitric oxide synthase, B-cell lymphoma-extra large, and protein expression of the endogenous nuclear factor-κB inhibitor). Although hyperoxia alone attenuated the postshock hyperinflammation and thereby tended to improve visceral organ function, hypothermia and combi treatment had no beneficial effect., Conclusions: During resuscitation from near-lethal hemorrhagic shock, hyperoxia attenuated hyperinflammation, and thereby showed a favorable trend toward improved organ function. The lacking efficacy of hypothermia was most likely due to more pronounced barrier dysfunction with vascular leakage-induced circulatory failure.

Background: This study aims to test the hypothesis whether lowering glycemia improves mitochondrial function and thereby attenuates apoptotic cell death during resuscitated murine septic shock., Methods: Immediately and 6 h after cecal ligation and puncture (CLP), mice randomly received either vehicle or the anti-diabetic drug EMD008 (100 μg · g(-1)). At 15 h post CLP, mice were anesthetized, mechanically ventilated, instrumented and rendered normo- or hyperglycemic (target glycemia 100 ± 20 and 180 ± 50 mg · dL(-1), respectively) by infusing stable, non-radioactive isotope-labeled (13)C6-glucose. Target hemodynamics was achieved by colloid fluid resuscitation and continuous i.v. noradrenaline, and mechanical ventilation was titrated according to blood gases and pulmonary compliance measurements. Gluconeogenesis and glucose oxidation were derived from blood and expiratory glucose and (13)CO2 isotope enrichments, respectively; mathematical modeling allowed analyzing isotope data for glucose uptake as a function of glycemia. Postmortem liver tissue was analyzed for HO-1, AMPK, caspase-3, and Bax (western blotting) expression as well as for mitochondrial respiratory activity (high-resolution respirometry)., Results: Hyperglycemia lowered mitochondrial respiratory capacity; EMD008 treatment was associated with increased mitochondrial respiration. Hyperglycemia decreased AMPK phosphorylation, and EMD008 attenuated both this effect as well as the expression of activated caspase-3 and Bax. During hyperglycemia EMD008 increased HO-1 expression. During hyperglycemia, maximal mitochondrial oxidative phosphorylation rate was directly related to HO-1 expression, while it was unrelated to AMPK activation. According to the mathematical modeling, EMD008 increased the slope of glucose uptake plotted as a function of glycemia., Conclusions: During resuscitated, polymicrobial, murine septic shock, glycemic control either by reducing glucose infusion rates or EMD008 improved glucose uptake and thereby liver tissue mitochondrial respiratory activity. EMD008 effects were more pronounced during hyperglycemia and coincided with attenuated markers of apoptosis. The effects of glucose control were at least in part due to the up-regulation of HO-1 and activation of AMPK.